This study evaluated APTT clot waveform analysis using the ACL TOP analyzer and compared it to analysis using the MDA 180 analyzer. APTT tests were performed on both analyzers in patients with hemophilia A and B as well as normal individuals. The APTT clot reaction derivatives produced by the ACL TOP correlated significantly with the clot waveform parameters from the MDA 180. Both sets of parameters correlated significantly with factor VIII and IX levels. The ACL TOP analysis demonstrated high sensitivity and specificity in differentiating between normal and hemophilia patients.

1. Evaluation of APTT clot waveform analysis by IL ACL TOP®
in
patients with Haemophilia and comparison to clot waveform
analysis with MDA®
180
Aghighi S., Riddell A.F., Vijayakumar, E., Chowdary, P.
Katharine Dormandy Haemophilia Centre & Thrombosis Unit, The Royal Free Hospital, Pond Street, London NW3 2QG, U.K.
Introduction
•Optical data generated during clotting tests such as prothrombin time (PT)
and activated partial thromboplastin time (APTT) have been used to measure
kinetics of fibrin generation, and indirectly of thrombin generation in
haemophilia patients 1
.
• Formation of fibrin leads to a concomitant increase in the optical density of
the plasma specimen (or a decrease in light transmittance).
• Modern automated photo-optical coagulation analysers, collect optical data
over the entire course of clot formation and an optical profile, or clot
waveform, is mathematically processed by a software algorithm, to derive
parameters of coagulation velocity.
• Clot waveform analysis using light transmittance on MDA®
180 analysers
has previously been reported 1,2,3
.
•Recently, Solano et al. have studied the use of APTT derivative curves by
using absorbance values to create the clot reaction curve on ACL TOP®
analysers Instrumentation Laboratory (IL), USA) in different patient groups 4
.
Conclusion
•The APTT clot reaction derivatives, produced by the ACL TOP®
correlated
significantly with the parameters of clot waveform analysis on the MDA®
180.
•Similar to MDA clot waveform analysis, ACL TOP®
results showed a
significant correlation with FVIII/FIX levels.
•The high sensitivity and specificity of these tests make them a reliable
predictor of FVIII/FIX levels in haemophilia patients.
Aim
To investigate the use of clot reaction curve derivatives, based on light
absorbance on the ACL TOP®
coagulometer, and compare them with the light
transmittance generated clot waveform as performed on the MDA®
180
analyser, in a normal individual group and in patients with haemophilia A.
References
1. Braun, P.J., Givens, T.B., Stead, A.G., Beck, L.R., Gooch, S.A., Fischer, T.J. (1997). Properties of optical data from APTT and PT
assays. Thromb Haemost, 78, 1079-87.
2. Downey, C., Kasmir, R.. Toh, CH. (1997). Novel and diagnostically applicable information from optical waveform analysis of blood
coagulation in disseminated intravascular coagulation. British Journal of Haematology, 98(1), 68-73.
3. Shima, M., Matsumoto, T., Fukuda, K., Kubota, Y., Tanaka, I., Nishiya, K (2002). The utility of activated partial thromboplastin time (aptt)
clot waveform analysis in the investigation of hemophilia a patients with very low levels of factor viii activity (FVIII:C). Thromb Haemost,
87(3), 436-441.
4. Solano, C., Zerafa, P., Bird, R. (2010). A study of atypical APTT derivative curves on the ACL TOP coagulation analyser. Int Jnl Lab Hem
33(1), 67-78.
Methods
•APTTs were measured on the MDA®
180 (Organon Teknika, UK), using
Platelin LS reagent (Stago, France) and on an ACL TOP®
, using SynthASIL
reagent (IL, USA). MDA®
180 clot waveform parameters (Min1 = velocity, Min2
= acceleration) and ACL TOP®
clot profile parameters (Max1 = velocity, Max2
= acceleration) were calculated by the manufacturer’s software.
•A reference range was established for each parameter and compared with 70
patients with Haemophilia (Haemophilia A: severe, n=27, moderate n=9, mild
n=10, and Haemophilia B: severe n=9, moderate n=3, mild n=4). FVIII and FIX
activity levels were also measured with one-stage clotting factor assay using
the ACL®
3000 analyser.
Results
•Min1 correlated significantly with Max1 (r = 0.883, p < 0.0001)
•Min2 also correlated significantly with Max2 (r = 0.883, p < 0.0001)
•FVIII levels correlated significantly with Min1, Min2, Max1, and Max2 (r =
0.804, 0.837, 0.701, 0.75;respectivly; p < 0.0001)
•FIX levels were also correlated significantly with Min1, Min2, Max1, and
Max2 (r = 0.721, 0.697, 0.685, 0.681 respectively; p < 0.001)
•Sensitivity and specificity of tests to distinguish between normal individuals
and patients with Haemophilia A or B was Min1 = 99%, 96%, Min2 = 97%,
96%, Max1 = 98%, 96% and Max2 = 100%, 96%.
Discussion
•Clot waveform analysis, based on light transmission on the MDA®
180
analyser, has been used to analyse fibrin clot formation beyond clot detection
as a surrogate marker of thrombin generation.
•Several groups have described the usefulness of clot waveform analysis in
different clinical settings 1,2,3
.
•In our study light absorbance based APTT derivatives generated by the ACL
TOP®
, correlated significantly with parameters of clot waveform by the MDA®
180. The high sensitivity and specificity we achieved make them a useful
global screening test.
•Our results showed a significant correlation between FVIII/FIX clotting activity
levels and Max1/Max2.
•APTT velocity and acceleration parameters of fibrin polymerisation may not
only be dependent on clotting factor levels but also other markers that may
influence the coagulation potential of individuals.
•Further studies are required to determine the correlation of APTT derivatives
and clinical phenotype of haemophilia patients.
Figure A showing linear regression
analysis of comparison of ACL TOP®
Max1 and MDA ®
180 Min1 (r2
=
0.771).
Figure B showing linear regression
analysis of ACL TOP®
Max2 and
MDA ®
180 Min2 (r2
= 0.789).
0 50 100 150
0
10
20
30
40
50
Min2
%
Max2
%
0 50 100 150
0
20
40
60
80
Min1
%
Max1
%
APTT clot reaction curve by ACL
TOP®
.
Based on light absorbance, point a is the
start of light absorbance by addition of
activating reagent. Point b indicates the
initiation of coagulation. Point c is the
coagulation midpoint. Point d is the end of
coagulation phase. Point e is the end of
acquisition time. The absolute value of
Max1 shows the maximum coagulation
velocity and the absolute value of Max2
shows the maximum coagulation
acceleration 4
.
APTT clot reaction curve by MDA®
180.
Based on light transmittance, point a is the
start of light transmittance by addition of
activating reagent. Point b indicates the
initiation of coagulation. Point c is the
coagulation midpoint. Point d is the end of
coagulation phase. Point e is the end of
acquisition time. The absolute value of Min1
shows the maximum coagulation velocity
and the absolute value of Min2 shows the
maximum coagulation acceleration 1
.
Max2
Max1
d
c
ba
mAbs
Time (s)
e
P-TH-568
Figure A Figure B
Min2
%
Min1
%